We propose to synthesize potent, stable and bioavailable opioid peptide analogues designed to be i) antagonists or inverse agonists with high selectivity for mu-, delta- or Kappa-opioid receptors and their subtypes as pharmacological tools and ii) opioid compounds with mixed agonist/antagonist profiles for possible therapeutic applications. To reach these goals, we use an interdisciplinary approach incorporating organic synthesis, peptide chemistry, extensive pharmacological characterization and conformational studies. Specific Aim 1 is based on our recent discovery that substitution of 3-(2,6-dimethyl-4-hydroxyphenyl)propanoic acid (Dhp) for the N-terminal tyrosine residue in opioid peptides produces antagonists at mu-, delta- or Kappa-receptors. We will synthesize Dhp analogues for Tyr1 replacement in opioid peptides that are capable of engaging in a variety of potential hydrophobic or hydrogen-bonding interactions with receptor moieties, with the expectation of obtaining more potent antagonists or inverse agonists, or compounds with a mixed K agonist/u antagonist profile. In Specific Aim 2 we will substitute the Dhp analogues developed in Specific Aim 1 for Tyr1 in known selective Kappa-, delta- or mu-agonist opioid peptides to convert them into antagonists or inverse agonists with selectivity for Kappa-, delta- or mu-receptors and their subtypes. In Specific Aim 3 we will develop potent K opioid antagonists or inverse agonists through structural modification of the cyclic tetrapeptide c[Phe-D-Pro-Phe-(D,L)-Trp] (CJ-15,208) recently isolated from the fungus Ctenomyces serratus. In Specific Aim 4 we will identify opioid peptide analogues containing certain Dhp analogues developed under Specific Aim 1 that have a mixed K agonist/mu antagonist profile as potential agents for the treatment of cocaine abuse. Specific Aim 5 concerns the development of chimeric opioid peptides with a mixed mu agonist/delta antagonist profile as analgesics with a low propensity to produce tolerance and dependence. The in vitro opioid activity profiles will be determined by performing opioid receptor binding assays, the guinea pig ileum and mouse vas deferens assays, and the [35S]GTPgammaS binding assay. The characterization of some of the compounds in vivo will include determination of the effects on cocaine discrimination and self-administration for the K agonist/mu antagonists and analgesic testing, including tolerance and dependence development, for the mixed mu agonist/delta antagonists. The conformation(s) of select compounds will be examined by 1HNMRspectroscopy in conjunction with theoretical conformational analyses.